<p>Excess excitation energy, mechanical injury and defence against pathogens, each trigger rapid production of reactive oxygen species (ROS) in <i>Arabidopsis thaliana</i> leaves. ROS, such as hydrogen peroxide (H₂O₂), are required for the induction of systemic acquired acclimation and may lead to redox changes in photosynthetic electron transport (PET). On one hand, enhanced ROS production during stress can destroy cells, and on the other, ROS can also act as signals for the activation of stress responsive and defensive pathways.</p><p>In this work, physiological and molecular analyses of <i>Arabidopsis</i> mutants and transgenic lines were applied to investigate the signalling network controlling biotic and abiotic stress responses. A key enzyme of the antioxidant network is encoded by <i>ASCORBATE PEROXIDASE 2 (APX2</i>). Wounded leaves showed low induction of<i> APX2 </i>expression and when exposed to excess light, <i>APX2</i> expression was increased synergistically. Signalling pathways dependent upon jasmonic acid, chitosan and abscisic acid were not involved in the wound-induced expression of <i>APX2</i>, but PET was required, and APX2 induction was preceded by a depressed rate of CO₂fixation.</p><p>Analysis of<i> lsd1</i> (<i>LESION SIMULATING DISEASE 1</i>) strongly suggests that light acclimatory processes and pathogen defences are genetically and functionally linked. It is important to know that LSD1 type of mutants have mainly been studied with regard to pathogenesis. From this work, it reveals that association of LSD1 with hypersensitive response may only be supplementary.</p><p><i>GLUTATHIONE PEROXIDASES</i> <i>(GPXs)</i> are another major family of ROS scavenging enzymes. Analysis of the <i>Arabidopsis </i>genome database revealed a new open-reading frame, thus increasing the total number of <i>AtGPX </i>gene family to eight (<i>AtGPX1-AtGPX8</i>). <i>Arabidopsis thaliana</i> transgenic lines with reduced expression of both putative chloroplastic isoforms (<i>AtGPX1 </i>and <i>AtGPX7</i>) and <i>AtGPX7</i> knock-out mutant (ko-<i>GPX7</i>) were more sensitive to photo-oxidative stress but had a reduced bacterial growth rate when inoculated with virulent strains <i>Pseudomonas syringae</i> pv. <i>tomato</i> DC3000 and<i> P.s.t. maculicola</i> strain ES4326, indicating increased resistance to pathogenesis. This, to our knowledge, is the first functional and genetic analysis of chloroplastic GPXs in plants, and confirms that light and chloroplastic ROS metabolism is important for basal resistance against virulent pathogens.</p><p>The above results confirm that light sensing, light acclimatory processes and photo-produced ROS also govern pathogen defence pathways. This has a great ecological relevance for Darwinian fitness of plants growing in the natural environment, where simultaneous pathogen attack and fluctuations in light, temperature and other environmental factors make rapid acclimation a constant necessity. Molecular, biochemical and physiological analysis of pathogen responses in mutants impaired in light sensing, EEE-dissipatory mechanisms, and similar analysis of light acclimatory processes in mutants impaired in pathogen defences may prove to be seminal.</p>

Excess excitation energy, mechanical injury and defence against pathogens, each trigger rapid production of reactive oxygen species (ROS) in Arabidopsis thaliana leaves. ROS, such as hydrogen peroxide (H2O2), are required for the induction of systemic acquired acclimation and may lead to redox changes in photosynthetic electron transport (PET). On one hand, enhanced ROS production during stress can destroy cells, and on the other, ROS can also act as signals for the activation of stress responsive and defensive pathways. In this work, physiological and molecular analyses of Arabidopsis mutants and transgenic lines were applied to investigate the signalling network controlling biotic and abiotic stress responses. A key enzyme of the antioxidant network is encoded by ASCORBATE PEROXIDASE 2 (APX2). Wounded leaves showed low induction of APX2 expression and when exposed to excess light, APX2 expression was increased synergistically. Signalling pathways dependent upon jasmonic acid, chitosan and abscisic acid were not involved in the wound-induced expression of APX2, but PET was required, and APX2 induction was preceded by a depressed rate of CO2 fixation. Analysis of lsd1 (LESION SIMULATING DISEASE 1) strongly suggests that light acclimatory processes and pathogen defences are genetically and functionally linked. It is important to know that LSD1 type of mutants have mainly been studied with regard to pathogenesis. From this work, it reveals that association of LSD1 with hypersensitive response may only be supplementary. GLUTATHIONE PEROXIDASES (GPXs) are another major family of ROS scavenging enzymes. Analysis of the Arabidopsis genome database revealed a new open-reading frame, thus increasing the total number of AtGPX gene family to eight (AtGPX1-AtGPX8). Arabidopsis thaliana transgenic lines with reduced expression of both putative chloroplastic isoforms (AtGPX1 and AtGPX7) and AtGPX7 knock-out mutant (ko-GPX7) were more sensitive to photo-oxidative stress but had a reduced bacterial growth rate when inoculated with virulent strains Pseudomonas syringae pv. tomato DC3000 and P.s.t. maculicola strain ES4326, indicating increased resistance to pathogenesis. This, to our knowledge, is the first functional and genetic analysis of chloroplastic GPXs in plants, and confirms that light and chloroplastic ROS metabolism is important for basal resistance against virulent pathogens. The above results confirm that light sensing, light acclimatory processes and photo-produced ROS also govern pathogen defence pathways. This has a great ecological relevance for Darwinian fitness of plants growing in the natural environment, where simultaneous pathogen attack and fluctuations in light, temperature and other environmental factors make rapid acclimation a constant necessity. Molecular, biochemical and physiological analysis of pathogen responses in mutants impaired in light sensing, EEE-dissipatory mechanisms, and similar analysis of light acclimatory processes in mutants impaired in pathogen defences may prove to be seminal.

Submitted by MARCELLO BONTEMPI PIZZI null (marcello.pizzi@gmail.com) on 2016-09-30T19:50:12Z No. of bitstreams: 1 Marcello_Bontempi_Pizzi_Dissertacao_UNESP_Desfolhas_Sucessivas.pdf: 7157176 bytes, checksum: 6cac889ca8ee38013ebd76f802772f05 (MD5) / Approved for entry into archive by Ana Paula Grisoto (grisotoana@reitoria.unesp.br) on 2016-10-04T17:29:25Z (GMT) No. of bitstreams: 1 pizzi_mb_me_bot.pdf: 7157176 bytes, checksum: 6cac889ca8ee38013ebd76f802772f05 (MD5) / Made available in DSpace on 2016-10-04T17:29:25Z (GMT). No. of bitstreams: 1 pizzi_mb_me_bot.pdf: 7157176 bytes, checksum: 6cac889ca8ee38013ebd76f802772f05 (MD5) Previous issue date: 2016-08-09 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A produtividade de plantios de eucalipto no Brasil foi triplicada nas últimas quatro décadas graças, principalmente, à pesquisa intensiva, investimentos em tecnologia silvicultural, clonagem e melhoramento genético. Atualmente, existem diversas pesquisas relacionadas ao gênero Eucalyptus, mas são poucas as que abordam o tema estresse biótico por ser uma avaliação complexa e de difícil planejamento e execução. Em Piracicaba, em um sítio experimental com alta sanidade e sobrevivência, foram induzidas uma, duas e três desfolhas sucessivas, retirando-se manualmente todas as folhas da copa das árvores de 12 clones de eucalipto de extrema relevância à silvicultura brasileira. As desfolhas ocorreram aproximadamente aos 15, 19 e 23 meses de idade das árvores, e a avaliação final de perda de produtividade ocorreu aos 27 meses. O crescimento das árvores foi monitorado semanalmente, sendo possível ver o efeito das desfolhas ao longo do período. Os clones de eucalipto escolhidos para este estudo, apesar de pertencerem ao mesmo gênero, mostraram resultados bastante diferentes. O impacto das desfolhas no crescimento das árvores foi imediato e prolongado, com o crescimento próximo à zero por 50 a 120 dias após a desfolha. As desfolhas causaram redução de crescimento em DAP (Diâmetro à Altura do Peito), altura total, volume e biomassa de madeira. As árvores não desfolhadas acabaram dominando as desfolhadas. Isso causou o estiolamento das árvores desfolhadas e a redução da homogeneidade das parcelas. Aos 27 meses, as árvores que sofreram uma desfolha apresentaram biomassa de madeira 48% inferior ao das árvores que não sofreram desfolhas (variando entre 24 e 57% para clones). Duas desfolhas sucessivas causaram redução média de 59% de biomassa (variando entre 49 e 72%) e três desfolhas sucessivas causaram perdas médias de 68% (variando entre 57 e 80%). / The Brazilian Eucalyptus productivity has tripled in the last four decades, mainly because of intensive research, development of forestry and genetics. Nowadays, there are several studies related to the Eucalyptus genus, but only a few address biotic stress, as the subject is of complex evaluation, difficult planning and execution. In Piracicaba, in an experimental site with high health and survival rate, one, two and three successive defoliations were induced, manually removing all the canopy leaves of 12 extremely relevant clones to Brazilian forestry. The defoliations occurred in approximately 15, 19 and 23 months after planting and the final evaluation of productivity occurred at 27 months. Tree growth was weekly monitored, and the effect of defoliation over time was visible. The eucalypts clones chosen for this study, despite belonging to the same genus, showed quite different results. The impact of canopy defoliation on tree growth was immediate and prolonged, with growth close to zero for 50 to 120 days following defoliation. Defoliation caused growth reduction in DBH (Diameter at Breast Height), total height, volume and wood biomass. Defoliated trees were etiolated as the non-defoliated were the dominant trees in the plot. Defoliation reduced plot homogeneity. One year after the first defoliation, the 27-month-old trees that have undergone one defoliation showed 48% lower wood biomass, in comparison with non-defoliated trees (ranging from 24 to 57% by clones). Two successive defoliations caused an average reduction of 59% of biomass (ranging from 49 to 72%) and three successive defoliations caused 68% of losses (ranging from 57 to 80%).

Submitted by THIAGO MATEUS ROSA DOS SANTOS null (thiagomateusrp@gmail.com) on 2017-07-28T16:24:35Z No. of bitstreams: 1 Dissertação_Thiago_Mateus_Rosa_Santos.pdf: 2449824 bytes, checksum: 8934ae7010672e8bf2342056d81d2899 (MD5) / Approved for entry into archive by LUIZA DE MENEZES ROMANETTO (luizamenezes@reitoria.unesp.br) on 2017-08-01T20:00:27Z (GMT) No. of bitstreams: 1 rosasantos_tm_me_jabo.pdf: 2449824 bytes, checksum: 8934ae7010672e8bf2342056d81d2899 (MD5) / Made available in DSpace on 2017-08-01T20:00:27Z (GMT). No. of bitstreams: 1 rosasantos_tm_me_jabo.pdf: 2449824 bytes, checksum: 8934ae7010672e8bf2342056d81d2899 (MD5) Previous issue date: 2017-05-03 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / A cana-de-açúcar é acometida por uma doença conhecida por “escaldadura das folhas” causada pela bactéria colonizadora do xilema Xanthomonas albilineans, considerada uma das principais doenças que atingem a cultura da canade-açúcar. A sintomatologia na fase crônica se caracteriza, principalmente, pelo aparecimento de uma faixa branca ao lado da nervura central da folha, a qual evolui para clorose total causando a morte da planta. Uma vez que o patógeno pode ser transmitido de várias maneiras, o seu controle demanda altos custos. Desta maneira, o desenvolvimento de cultivares tolerantes é uma boa opção para o controle efetivo da doença. A tolerância e sensibilidade das plantas aos fatores bióticos está relacionada com a expressão de genes, e dentre estes, os miRNAs (incluindo o miR397 e o miR319) têm sido relatados como importantes reguladores em vários mecanismos de resposta das plantas. O objetivo deste trabalho foi analisar a expressão de dois miRNAs (miR319 e miR397) em duas cultivares de cana-de-açúcar (RB86-7515 - tolerante e SP78-4467 - suscetível), infectadas por uma linhagem de X. albilineans (IACXa11), considerada a mais virulenta do Brasil. Para isto, as plantas foram cultivadas em vasos, inoculadas com X. albilineans e mantidas em casa de vegetação. Amostras de folhas e colmos foram coletadas em cinco períodos (24, 72, 144, 360 e 720 h) e a expressão dos miRNAs foi analisada pela técnica de Stem-loop RT-qPCR. Os miR397 e miR319 apresentaram-se diferencialmente expressos nas cultivares e entre os tecidos. Na cultivar suscetível (SP78-4467), durante os primeiros períodos de infecção (24, 72 e 144 h), houve uma resposta tardia de defesa quando comparada com a cultivar tolerante (RB86-7515). O miR319 apresentou o mesmo perfil de expressão em folhas e colmos da cultivar RB86-7515 (tolerante), sugerindo que o reconhecimento do patógeno e a ativação dos mecanismos de defesa são modulados em ambos os tecidos. De maneira geral, as análises dos miRNAs demonstraram que a expressão do miR397 é menor quando comparada com o miR319. O mesmo padrão foi observado para os seus respectivos genes alvo. O miR397 regula a enzima lacase, importante na biossíntese de lignina. A repressão deste miRNA aumentaria a lignificação, sugerindo um mecanismo estrutural de resposta. O miR319 regula os fatores de transcrição (FTs) MYB e TCP, os quais são responsáveis pela sinalização de ácido abscísico (ABA) e ácido jasmônico (JA). A repressão destes hormônios vegetais desencadeia a sinalização por ácido salicílico (SA), o qual é responsável pela defesa contra patógenos hemibiotróficos, tal como X. albilineans. / Sugarcane is affected by a disease known as "leaf scald" caused by the bacterium Xanthomonas albilineans, which colonizes the xylem. This disease is one of the most important for sugarcane culture. The chronic phase is mainly characterized by the white band emergence along the central leaf vein, which causes total chlorosis of the leaf and plant death. Since the pathogen can be transmitted in many ways, his control demands high costs, and the development of tolerant cultivars is a good option for disease control. The plant tolerance and sensitivity to biotic factors is related to gene expression, and among these, the miRNAs (including miR397 and miR319), have been reported as important regulators in various plant response mechanisms. The aim of this work was to analyze the expression of two miRNAs (miR319 and miR397) in two sugarcane cultivars (RB86-7515 – tolerant, and SP78- 4467 - susceptible), infected by a strain of X. albilineans (IACXa11), the most virulent in Brazil. The plants were grown in vases, inoculated with X. albilineans and kept in a greenhouse. Samples of leaves and stems were collected in five periods (24, 72, 144, 360, and 720 h), and the miRNA expression was analyzed by Stem-loop RT-qPCR. The miR397 and miR319 expression were different between cultivars and tissues. In the susceptible cultivar (SP78-4467), during the first infection periods (24, 72 and 144 h), there was a late defense response when compared to the tolerant cultivar (RB86-7515). The miR319 presented the same expression profile in leaves and stems of the cultivar RB86-7515 (tolerant), suggesting that the pathogen recognition and defense mechanisms activation were modulated in both tissues. In general, miRNAs analyzes demonstrated that miR397 expression is lower when compared to miR319. The same pattern was observed for their respective target genes. The miR397 is a laccase regulator, important in lignin biosynthesis. Repression of this miRNA would increase lignification, suggesting a structural mechanism of response. The miR319 regulates the transcription factors (TFs) MYB and TCP, which are responsible for the abscisic acid (ABA) and jasmonic acid (JA) signaling. The repression of these plant hormones triggers salicylic acid (SA) signaling pathway, which is responsible for the defense against hemibiotrophic pathogens, such as X. albilineans. / CNPq: 153785/2014-4

Magister Scientiae - MSc / Stress responsive genes play a key role in shaping the manner in which plants process and respond to environmental stress. Their gene products are linked to DNA transcription and its consequent translation into a response product. However, whilst these genes play a significant role in manufacturing responses to stressful stimuli, transcription factors coordinate access to these genes, specifically by accessing a gene's promoter region which houses transcription factor binding sites. Here transcriptional elements play a key role in mediating responses to environmental stress where each transcription factor binding site may constitute a potential response to a stress signal. Arabidopsis thaliana, a model organism, can be used to identify the mechanism of how transcription factors shape a plant's survival in a stressful environment. Whilst there are numerous plant stress research groups, globally there is a shortage of publicly available stress responsive gene databases. In addition a number of previous databases such as the Generation Challenge Programme's comparative plant stressresponsive gene catalogue, Stresslink and DRASTIC have become defunct whilst others have stagnated. There is currently a single Arabidopsis thaliana stress response database called STIFDB which was launched in 2008 and only covers abiotic stresses as handled by major abiotic stress responsive transcription factor families. Its data was sourced from microarray expression databases, contains numerous omissions as well as numerous erroneous entries and has not been updated since its inception.The Dragon Arabidopsis Stress Transcription Factor database (DASTF) was developed in response to the current lack of stress response gene resources. A total of 2333 entries were downloaded from SWISSPROT, manually curated and imported into DASTF. The entries represent 424 transcription factor families. Each entry has a corresponding SWISSPROT, ENTREZ GENBANK and TAIR accession number. The 5' untranslated regions (UTR) of 417 families were scanned against TRANSFAC's binding site catalogue to identify binding sites. The relational database consists of two tables, namely a transcription factor table and a transcription factor family table called DASTF_TF and TF_Family respectively. Using a two-tier client-server architecture, a webserver was built with PHP, APACHE and MYSQL and the data was loaded into these tables with a PYTHON script. The DASTF database contains 60 entries which correspond to biotic stress and 167 correspond to abiotic stress while 2106 respond to biotic and/or abiotic stress. Users can search the database using text, family, chromosome and stress type search options. Online tools have been integrated into the DASTF, database, such as HMMER, CLUSTALW, BLAST and HYDROCALCULATOR. User's can upload sequences to identify which transcription factor family their sequences belong to by using HMMER. The website can be accessed at http://apps.sanbi.ac.za/dastf/ and two updates per year are envisaged. / South Africa

<p>  </p> <p>Modern agriculture must address the massive challenge of providing food for the increasing population. The challenge lies in increasing crop yield and reducing losses caused by abiotic and biotic stresses. In fact, for some crops, such as wheat and maize, over 40% of the production is lost due to environmental conditions (abiotic stresses) or pests and pathogens (biotic stresses). Specialists in the area are suggesting a need for a second green revolution to meet the increasing demand in food production. While in the first green revolution was focused on breeding and genetics to produce crops' genetic lines with a higher yield. The second green revolution will utilize cutting-edge technologies to increase yield and reduce crop losses. The development of remote sensing technologies and their applications is the main driving force of modern agricultural practices. Currently, farmers are relying more on automation, data collection, and data analysis to manage farming operations. The reliance on remote sensor technologies is a game-changer for traditional agricultural practices, and it is contributing tremendously to increasing production and avoiding yield losses. Hyperspectral phenotyping is an emerging remote sensing technology that utilizes the light's reflectance to provide insightful information about plant traits. For several years, research groups have been applying hyperspectral phenotyping techniques to detect plant traits information, such as nitrogen content, photosynthesis rates, pests infestation, and abiotic stress detection. Although this is not a novel approach to plant traits detection, this technology application is not mature yet. Several challenges are associated with using hyperspectral information for phenotyping, such as model transferability, data collection scalability, and the heritability of plant traits retrieved using hyperspectral data. In my thesis dissertation, I addressed some of those challenges contributing to advances in hyperspectral phenotyping. My results demonstrate that using full-range hyperspectral reflectance data (400-2400nm) to retrieve nitrogen in winter wheat increases the model transferability across years and genotypes. Predicting nitrogen content using hyperspectral data can be used as a surrogate to calculate nitrogen use efficiency traits. My research highlights the hurdles associated with spectral detection of stresses interaction, such as drought stress, which can mask western corn rootworm detection in maize. Finally, I explored the correlation among spectral, functional, and field traits in a soybean NAM (Nested Association Mapping) population to understand the relationship among those traits' variability and how that information can be used for soybean breeding programs. The outcomes of my thesis dissertation advance the knowledge in the hyperspectral phenotyping field and its application to modern agriculture. Consequently, my study also contributes to food security programs by providing insightful information about the hyperspectral assessment of plant health status, which is essential to increase yield production and reduce crop losses. </p>

Biotic and abiotic stresses are signifi cant factors limiting the production of food and other supporting materials required to sustain increasing world population. Plant health is directly related to human health and is increasingly becoming signifi cant and demands more attention towards limiting the damages caused by biotic and abiotic stresses. Signifi cant progress has been made towards our understanding of the processes, which mediate both biotic and abiotic stress signaling in plants. Signifi cant role is played by various plant hormones, e.g., salicylic acid (SA) and jasmonic acid (JA) in biotic stress and abscisic acid (ABA) in abiotic stress (Annu Rev Cell Dev Biol 28:489-521, 2012). Other hormones with minor role include the cytokinins (CK), auxins (indole 3 acetic acid. IAA), and the brassinosteroids (BR) (Annu Rev Cell Dev Biol 28:489-521, 2012). Cross talk between these plant hormones is signifi cant and may result in either synergistic or antagonistic effect on stress responses (Annu Rev Cell Dev Biol 28:489-521, 2012). In recent years, extensive research carried out in various laboratories has implicated cross talk between the ABA and the SA in abiotic stress response. This is signifi cant in light of SA being key player in biotic stress responses in plants. This review will discuss the role of SA in biotic and abiotic stress signaling and its cross talk with other hormones in mediating abiotic stress signaling in plants.

The Leaf Economics Spectrum (LES) describes the continuous range of leaf ecophysiological strategies that plants across the globe utilize to achieve a net-positive return on their resource investment. This spectrum is measured by traits such as leaf mass per area, leaf nitrogen, and leaf lifespan. Unsuitable leaf trait combinations are selected against in nature due to a net-loss on the return of initial resource investment, while some potentially extremely effective trait combinations may not be possible due to genetic constraints. Existing genetic variation in LES traits, however, suggests the spectrum may be less rigid than previously expected. To test this, we exploit phenotypic plasticity by subjecting cultivated sunflower, Helianthus annuus, to environmental stress to generate variability in LES traits. In a greenhouse experiment, treatment groups of Helianthus were exposed to six varying intensities each of four acute stress applications: nutrient limitation, high soil salinity, heat, and simulated herbivory. Leaves formed during the onset of stress and post-stress were sampled to measure leaf traits; changes in plant fitness were estimated through total plant biomass and the time of first flowering. Results from statistical analyses show a variety of significant effects that stress applications had on plant growth and LES traits. Our findings explore the possibility of achieving off-axis LES trait combinations through salt-induced trait plasticity, and if further studies are needed to reassess the rigidity of the spectrum.

Un des principaux acteurs impliqués dans la régulation du statut redox intracellulaire, permettant aux plantes de s’adapter aux contraintes environnementales, est une famille multigénique de petites (12-14 kDa) oxydoréductases ubiquistes appelées thiorédoxines (TRX). Le génome d’Arabidopsis code une vingtaine de TRX canoniques dont neuf sont plastidiales (TRX f, m, x et y). Très étudiées dans notre laboratoire par des approches biochimiques, les TRX de types f et m apparaissent réguler majoritairement l’activité d’enzymes impliquées dans le métabolisme primaire tandis que les types x et y servir principalement de substrats réducteurs d’enzymes antioxydantes. Plus récemment, une dixième TRX, proposée plastidiale et nommée TRX z, a été identifiée. Au cours de ma thèse, j’ai caractérisé cette nouvelle TRX chloroplastique montrant des propriétés physico-chimiques inhabituelles, la rendant unique. En effet, la TRX z semble interagir, via des interactions électrostatiques, avec des protéines pour former des complexes de masses moléculaires élevées, potentiellement liés aux acides nucléiques. De plus, la TRX z, dont l’expression est induite à la lumière, principalement dans les tissus photosynthétiques, est la première TRX chloroplastique qui n’est pas réduite par le système FTR à la lumière mais qui peut être réduite par les autres TRX plastidiales, suggérant une interconnexion entre ces différentes TRX. D’autre part, une recherche exhaustive de cibles de la TRX z, par deux approches spécifiques et complémentaires (protéomique et double hybride), ont révélé 90 cibles putatives de la TRX z. La plupart de ces cibles, jamais identifiées comme cibles des TRX, sont impliquées dans la réponse de défense des plantes mise en place lors de stress biotiques. Ces résultats suggèrent que la TRX z pourrait être un élément clé dans la mise en place de ces réponses. L’analyse fonctionnelle préliminaire de la TRX z au cours de la réponse immune innée conforte cette hypothèse. L’ensemble de ces résultats indique que la TRX z pourrait jouer le rôle d’une protéine senseur de l’état d’oxydoréduction de la cellule. / One of the main actors involved in regulation of the cellular redox state, which allow plant adaptation to stress environmental conditions, is a multigenic family of small (12-14 kDa) ubiquitous oxidoreductases named thioredoxins (TRX). Arabidopsis encodes around twenty canonical TRX, including nine plastidial isoforms (TRX f, m, x and y). Extensively studied in our laboratory by biochemical approaches, TRX f and m was found to mainly redox regulate the activity of enzymes involved in the primary metabolism whereas TRX x and y serve as reducing substrates for antioxidant enzymes. More recently, a tenth TRX, predicted plastidial and named TRX z, was identified. During my PhD, I have characterized this new plastidial TRX showing unusual physicochemical properties, making it unique. Indeed, TRX z seems to interact, via electrostatic bonds, with proteins to form high molecular weight complexes, potentially linked to nucleic acids. Moreover, TRX z, which is expressed in green tissues in the light, is the first plastidial TRX which is not reduced by the FTR system but which can be reduced by other plastidial TRX, suggesting an interconnection between these TRX. Furthermore, a large scale inventory of TRX z targets, by two specific and complementary approaches (proteomic and yeast two hybrid), revealed 90 putative TRX z targets. Most of these, which have never been identified as TRX targets before, are implicated in plant defense response to biotic stresses. These results suggest that TRX z might be a key player in these responses. Preliminary functional analysis of TRX z during immune innate response reinforces this hypothesis. Altogether, these results indicate that TRX z appears as an important sensor of the redox status of the cell.

Made available in DSpace on 2016-12-23T13:48:24Z (GMT). No. of bitstreams: 1 Silas_Rodrigues_Disseracao_Completa.pdf: 3107447 bytes, checksum: 7c277a205cc0673835ce714a89f3efd0 (MD5) Previous issue date: 2006-08-25 / A meleira do mamoeiro é causada pelo Papaya meleira virus (PMeV), de genoma de dsRNA, que induz uma exsudação espontânea de látex fluido à partir de frutos e folhas. Inicialmente, objetivou-se estabelecer uma metodologia molecular simplificada para o diagnóstico precoce do vírus. A metodologia baseou-se na extração e análise do dsRNA viral, a partir de látex e tecidos da planta. A transmissão do vírus pela mosca-branca Trialeurodes variabilis foi avaliada, observando-se que o inseto não transmitiu o vírus, embora tenha sido capaz de adquirí-lo a partir de mamoeiros infectados. Foi confirmado que a infecção ocorre quando látex infectado é injetado no ápíce do caule do mamoeiro. A avaliação da distribuição do PMeV em mamoeiros em diferentes fases da infecção revelou a ocorrência do vírus, principalmente, nos órgãos ricos em laticíferos. Neste tipo de célula, o vírus reduziu a quantidade e alterou a morfologia sueprficial dos polímeros presentes no látex, ligando-se fortemente à estes polímeros. O PMeV induziu um acúmulo dos elementos potássio e fósforo no látex, que também apresentou teores mais elevados de água. Observou-se também no látex infectado a síntese de alcalóides e um acúmulo de cristais de oxalato de cálcio. Este acúmulo foi positivamente correlacionado com a síntese de H2O2 nos laticíferos de plantas doentes, que também foi observada nas células companheiras do floema. Os resultados obtidos sugerem a existência de um mecanismo de defesa do mamoeiro contra o PMeV que envolve a síntese de H2O2 pelos laticíferos e células do floema. / Papaya sticky disease' is caused by Papaya meleira virus (PMeV) which has a dsRNA molecule as genome. It induces an exudation of fluid latex from the fruits and leaves. Initially, this work aimed to establish an easy molecular diagnostic method to early virus detection. The stablished method was based on virus de dsRNA extraction from papaya latex and tissues. Afterwards, the PMeV transmission by Trialeurodes variabilis whitefly was evaluated. Our observation was that as far as the insect caried the virus, it did not transmit it to the papaya plant. We also established that PMev infection occurs only when latex from diseased plants was injected in healthy ones. The evaluation of PMeV allocation within papaya at different infecton stages demonstrated that the virus was present mainly in laticifers richly tissues. In this cell type, the virus strongly reduced the amount of latex polymers, also altering their superficial morphology. Using transmission elecron microscopy, PMeV was observed strongly linked to the polymers. The virus induced an ncrease in water and potassium and phosphorus elements levels in the latex. Also, it was observed an increase in alkaloids and calcium oxalate crystal accumation. This accumulation was positively correlated with H2O2 synthesis, which was also observed at phloem companion cells. Taken together, these results suggest the existence of papaya defense mechanism against PMeV involving the H2O2 synthesis by laticifers and phloem cells.